SE453191B - TIOALCANOYL CARNITINES, PROCEDURES FOR THEIR PREPARATION AND PHARMACEUTICAL COMPOSITIONS WITH MUCOLYTIC AND HOUSING DEMONSTRATIVE ACTIVITIES CONTAINING THEM - Google Patents

Description

453,191 hours! is in the terminal position, with a compound of the general formula III + 3)3N-CH2-fH-CH2 X- OCO(CH2)n~CH=CH2 (CH -COOH in which n is an integer between 0 and 4, according to the following reaction formula: + R COSH+(CH3)3N-CH -CH-CH -COOH-9(I, terminal thioalkanoyl) 1 2 2 x" oco(cH2)n-cH=cH2 I or (b) if in the compounds I the thioalkanoyl group R1COS- is in the non-terminal position, with a compound of the general formula IV A + c (CH3)3N~CH2-CH-CH2-COOH x" oco- (CHZ) mcH=cH- (c112)m1-R2 in which m and ml are each an integer between 0 and 3, ! and R is a lower alkyl group having 1 to 4 carbon atoms, 2 according to the following reaction formula: + R COSH+(CH3)3N-CH -CH-CH -COOH-?(I, non-terminal thioalkanoyl) l 1 2 2 X OCO(CH2)m~CH=CH-(CH2)m1-R2 The group R is preferably selected from the class comprising ethylene, trimethylene, tetramethylene, ethyl~ethylene, propylene, 1-methyl-trimethylene and 2-methyl-trimethylene.

The groups R1 and R2 are preferably selected from the class comprising methyl, ethyl, propyl, isopropyl and butyl.

The reaction between II and III and between II and IV takes place very easily. It is sufficient to bring the reactants into contact with each other even in the absence of solvent and suspending agent and to keep the reaction mixture under stirring from a few hours to a few days, at a temperature between about room temperature and 50°C. It is advantageous to use an excess of thioacid II in relation to .

III and IV. the excess varies from about 3:1 (molar) if the reaction is carried out in the presence of solvent to 30:1 (molar) in the absence of solvent.

Solvents and any suspending agents, as well as the isolation and purification measures, are those commonly used in organic synthesis work.

The following non-limiting examples illustrate the process for preparing some of the compounds of the invention and their chemical-physical properties.

Example 1 3-thioacetyl-propionyl-carnitine (ST 405) + + (CHB) 3-N-CH2-CH-CH2-COOH (CH3) 3-N-CH2-CH- CHz-COOH cl' oco(-cn2)2-Br cl' oco-cH=cH2 + (CH3)3-N-CH2-CH-CH2-COOH Cl OC0-(CH2)2-SCO-CH3 <1) Ezëæëëëllnia9_§y_§ëzxl9xl:§ëraiëia A solution of 4 g bromopropionyl-carnitine in 40 ml H20 was added with 100 ml IR 45-amberlite resin, which had been activated in the OH form. The reaction mixture was kept under stirring for 2 hours, then filtered and lyophilized.

The crude material thus obtained was purified by chromatography on silica gel buffered with 1.5% Na2HPO4 and eluting with methanol. The eluate was treated with HCl and then lyophilized; 1.5 g of pure product was obtained. Yield 42%.

Nuclear magnetic resonance D 2 O 6 6.6 - 6.3 (3H, m, CH=CH 2 ): 5.6 (1H, m, -CH-); CH3 + + 3.8 (zu, m,šš>N-cn2); 3.3 (9H, S, cn3íš>N-); CH3 2.6 (zu, d, -cuzcoa. 455 191 (2) (CH3)3-ä-CH -CH-CH -COOH U) "in Egaparation of 3-thioacetyl-propionyl-carnitine Inner salt of acryloyl-carnitine (0.008 mol), obtained according to the previous reaction step (1), was suspended in 40 ml of absolute ethanol. Thioacetic acid (0.0238 mol) was added to the resulting suspension. The suspension was kept under stirring for 3 h, then precipitated with anhydrous diethyl ether and stored in a refrigerator overnight.

An oil precipitated, which was separated by decantation and purified repeatedly by dissolution in ethanol and precipitation with diethyl ether. The crude material thus obtained was taken up in water, acidified with concentrated hydrochloric acid at 0°C and then lyophilized. 1.9 g of product was obtained. Yield 75%.

Nuclear magnetic resonance D20 6 5.6 (1H, m, -fñ-); O + + 3,s (211, m,>N-CHZ-); 3.3 (en, s, (cH3)3-N); 3.0 - 2.6 (GH, m, -QÉZCOOH, -Cogëzggz-S-); 2.3 (3H, s, -COCH3).

Example 2 5-thioacetyl-pentanoyl-carnitine (ST 412) + .

(CH3)3-N-CH2-CH-CH2-COOH OCO"(CH2)2-CH=CH 2 l 2 cl' on "__-á 2 (CH3) -É-CH -CH-CH2~COOH 3 2 cl" oco- (CH -scocH 2)4 3 §ram§tällnig¶_av allyl-acetyl-carnitine To a solution of carnitine hydrochloride (9 g; 0.045 mol) in 15 ml trifluoroacetic acid was added 0.13 mol allyl-acetyl-chloride. The resulting solution was kept at 45°C for 4 h. Then acetone was added to the solution, unreacted carnitine was separated and diethyl ether was added, whereby a precipitate was obtained. The crude material thus obtained was purified by precipitation with isopropanol-diethyl ether. 9.5 g of product was obtained. Yield 66%.

Nuclear magnetic resonance D20 6 5.7 (2H, m, -§§= -fñ-); 0 cnz, 5.1 (za, m, -cH= ggz-); 3.8 (za, m,E3>§-CH2->; + 3.3 (sn, s, (ca -N), 2.8 (2H, a, -gg2coon); 3)3 2.5 (4H, m, -COCHZ-CH2-). <2) Eaëmsëëlaias_ay_â:§i9ëes§xl:e§a§ëa9xlzëëšaiëlæ 22.4 g (0.38 mol) thioacetic acid was added to the allyl-acetyl-carnitine (4 g; 0.013 mol) from the previous reaction step <1). The resulting solution was kept at 4o°c overnight. Diethyl ether was then added to the solution and the precipitate thus obtained was isolated by decantation. The precipitate was dissolved in water and extracted three times with diethyl ether. The aqueous phase was concentrated and washed with acetone, yielding 3.9 g of product. Yield 84%.

Nuclear magnetic resonance D20 6 5.6 (1H, m, -?H~); I O 3.7 (ZH, m[f 3 >N-CH 2 -); 3.2 (9H, s, (CH 3 ) 3 -N-); 2.7 (en, m, ¿gg2cooH; ococH2-; cH2s): 2.4 (3H, S, -COCH3); 1.6 (4H, m, ~C0CH2§§2§§2CH2S-). 455 191 (1) (2) Example 3 3-thioacetyl-butyryl-carnitine (ST 406) (cH3)3-É-CHZ-ca-cuz-coon + cH cH=cHcoc1 3 cl' on _ Å/ + (CH3)3-N~CH2-FH-CH2-COOH cl' ococH= CH-CH3 i + (CH3)3-N-CH2-ïH~CH2~COOH Cl OCOCH2~ïH-CH3 SCÖCH3 Eransäëllni§¶_§!_ë§9E9e9xl:äëzaiëie 8 ml (0.08 mol) crotonoyl chloride was added to a solution of carnitine hydrochloride (8 g; 0.04 mol) in trifluoroacetic acid. The reaction mixture was kept under stirring at 50°C overnight. Then the mixture was added with diethyl ether and a precipitate formed. The precipitate was filtered off and used as such for the subsequent reaction.

Nuclear magnetic resonance D20 6 7.5 - 6.9 (1H, m, -ocogg; cH-); 6.3 - 5.5 (2H, m, -ococH=gg-; -ene-); o + 3.8 (zu, mj§5>N-cnz-); 3.3 (en, s, (ca 351131; 2.8 (ZH, d, -§§2COOH); 2.0 (3H, d, =CH-ÉEB).

Ešaeëëëlleia9_ëy_§:§i9ë2§§xl:ëQ§y§1l:äëšaiiia 15 ml (0.20 mol) of thioacetic acid was added to the crotonyl carnitine (3.5 g; 0.01 mol) from the previous reaction step (1). The reaction mixture was kept under stirring at room temperature for 4 days. Diethyl ether was then added to the reaction mixture. An oil was obtained, which was purified by dissolution in ethanol and reprecipitation with diethyl ether. The precipitation was repeated three times. Then the precipitate was dissolved in water and extracted three times with diethyl ether. The aqueous solution was lyophilized. 2.6 g of product were obtained. Yield 70%.

Nuclear magnetic resonance D20 6 5.5 (1H, m, -?H-); O + + 3.8 (2H, m,§5>N-CH2-); 3.2 (9H, s, (CH 3 ) 3 -N); 2.6 - 2.9 (fm, m, -ggz coon; -ococnz-n 2.4 (3H, S, -COCH3); 2.2 (1H, m, -CH-SCO-); 1.4 (3H, d, -CH-Ega).

The acute toxicity, expectorant effect, mucolytic effect and effect on ciliary motility of the compounds of formula I were determined.

Acute toxicity LD50 for the compounds of formula I was determined by Weil's method ("Tables for convenient calculation of median effective dose (LD50 or ED50) and instructions in their use", Biometrics, pages 249-253, 1952) after intraperitoneal administration to mice and is shown in Table 1.

Table 1 LD 50 and confidence limits, mg/kg intraperitoneally, for the compounds of formula I according to Weil's method N=4, K=4.

Compound LD50 confidence limits ST-405 378 489 - 292 ST-406 97 115 - 82 ST-412 1540 1760 - 1320 453 191 Coughing effect ~ Tests were performed on male rabbits weighing 2-3 kg, anesthetized with ethyl urethane, using the following method described by Perry et al.

(J. Pharm. Exp. Ther. lie, 65, 1941).

The anesthetized animals, which were secured with their heads down on an operating table at an angle of 60°, had a cannula inserted into the trachea. Each cannula was connected to a feeding device which provided a steady stream of preheated air (36-38°C) with constant humidity (80%). At the lower end of each cannula was fitted a graduated cylinder in which the bronchial secretion was collected. All the animals breathed spontaneously and consequently the air intake was automatically regulated to that suitable for normal breathing. One hour after insertion of the cannula, the compounds of formula I were administered orally (by gastric tube) to the animals in doses between 20 and 40 mg. Each dose of the drug was administered to 5 animals. The control animals (8) received only water. The amount of secretion was determined 1, 2 and 4 hours after administration. The results, summarized in Table 2, show that the compounds of general formula I have no expectorant effect.

Mucolytic action The tests were carried out in vitro using the method of Morandini et al. (Lotta contro la tubercolosi 47, n. 4, 1977). A platelet counter was used to follow the variations that the compounds of formula I and acetylcysteine induce on the rheological properties of human sputum. The results summarized in Table 3 show that the compounds of formula I produce a greater reduction in the density of human sputum than that induced by acetylcysteine.

Effect on ciliary activity The ability of the compounds of formula I to affect ciliary motility was studied microscopically on rat tracheal rings in solutions of the test compounds.

U Through this technique, it is possible to study, with reference to the concentration and contact time of the compounds, the blockage of ciliary movement, which is caused by the test compounds 453,191 and can be attributed to mucus secretion from the ciliary epithelium.

Substances to be used in the form of solutions must allow the indicated blocking to take place within not less than fifteen minutes after contact. 2% solutions of the compounds of formula I caused the blocking of ciliary movement to take place within 18-20 minutes.

Table 2 Effect of compounds of general formula I on bronchial secretion Percentage variations 1 standard error in bronchial secretion in relation to baseline values at the following time intervals Compounds after administration 1 hour 2 hours 3 hours Control (H20) +1.3x0.04 +2.210.05 +3.5æ0.04 ST-405 +1.1:0.03 +1.910.04 +3.6i0.05 ST-406 +1.410.05 +1.910.05 +3.4l0.04 ST-412 +1.3:0.04 +2.1:0.05 +3.1:0.03 n = 6 animals in each group 453 191 Table 3 U Mucolytic effect in vitro of compounds of formula I and acetylcysteine; change in density of human sputum Decrease in % 1 standard error of the line in relation to the peak of the maximum (*) after addition of 1 ml of a 10% solution of the test substance at the indicated dilution Compounds 1/30 1/60 ST-405 79 65 ST-406 88 70 ST-412 87 64 acetylcysteine 84 55 (*) mucolytic activity index As experimentally demonstrated, the compounds according to the invention clearly modify the rheological properties of sputum. When examining the results obtained, a decrease in sputum density was observed at higher doses (or lower dilutions) and at lower doses (or higher dilutions) consistently higher than that induced by acetylcysteine. On the other hand, neither of the compounds that increase bronchial secretion is capable of blocking the ciliary movement in the epithelium of rings of a trachea within shorter time intervals than those permitted.

The compounds of the present invention can be used therapeutically for the treatment of respiratory diseases.

A therapeutically effective amount of an ester of the general formula I is administered orally or parenterally to patients in need thereof.

The orally or parenterally administered dose of the compound of general formula I is generally between about 15 and about 70 mg/kg body weight per day, although larger or smaller doses may be administered by the supervising physician 453 191 11 taking into account age, weight and general condition, using proven medical experience.

In practice, the compounds are administered orally or parenterally in any of the usual pharmaceutical forms, which are prepared by methods well known to those skilled in the art of pharmaceutical technology. These forms are solid and liquid oral unit doses, e.g., tablets, capsules, solutions, syrups, and the like, and injectable forms, such as sterile solutions for ampoules and vials.

Some non-limiting examples of compositions suitable for oral or parenteral administration are set forth below.

Pharmaceutical compositions Ampoules for aerosol administration or intramuscular administration Each ampoule contains: ST-405 0.40 g sodium metabisulfite 10 mg pyrogen-free, distilled water 3 ml sleeve: ST-405 4.0 g sorbitol, 70% 15 g sucrose 50 g ethanol 1 ml p-hydroxybenzoate 0.2 mg flavorings 0.5 ml distilled water q.s. to 100 ml saccharin 0.20 9 453 191 12 Suppositories for adults ST-405 sodium metabisulfite excipient q.s. for 1 suppository Suppositories for children ST-405 sodium metabisulfite excipient q.s. for 1 suppository 0.40 o,o2o Suppositories for children still nursing ST-405 sodium metabisulfite excipient q.s. for 1 suppository Qoseringspoffe (5 Q) 100 g each contains: ST-405 saccharin flavoring orange freeze-dried orange juice sucrose, the rest up to 100 g 0.101 0.005 3.80 0.20 o,s 10 l-Q LQLQKQUJ (I

Claims (1)

1. 0 15 20 25 30 35 where 2. reach. 5. that 453 191 n I: CLAIMS Thioahaumyl carnitines of the general formula I + (cH 3) 3 N - CH 2 - en - CH 2 - coon X 'oco - R - sco - R 1 R is a straight alkylene group having 2 to 6 carbon atoms or a branched alkylene group having 3 to 6 carbon atoms, R 1 a straight or branched lower alkyl group having 1 to 4 carbon atoms and X "a halide ion. Thioalkanoyl carnitine according to claim 1, characterized in that it consists of 3-thioacetyl-propionyl thioalkanoyl-carnitine according to Claim 1, characterized in that it is 5-thioacetyl-pentanoyl-carnitine Thioalkanoyl-carnitine according to Claim 1, characterized in that it consists of 3-thioacetyl-butyryl Process for the preparation of thioalkanoylcarnitines of the general formula I + (cH3) 3N - caz - en - cH2 - coon x 'oco - R - sco - R 1 R is a straight alkylene group having 2 to 6 carbon atoms or a branched alkylene group having 3 to 6 carbon atoms, R a straight or branched lower alkyl group having 1 to 4 carbon atoms and X a halide ion, according to claim 1, I5 453 1 91 It is characterized in that (a) in case the azio compounds of formula I thioalkanoyl group R1COS- are in the terminal position, a compound of general formula III + (cH3) 3N ~ cH2 - cH -, cH - coon 2 X 'oco (cH2) n - CH = cnz where n is an integer between 0 and 4, is reacted with a thioacid of the general formula II R cosn 1 where R1 has the meaning given above, as follows, reaction formula R1cosn + (cH3) 3ñ - cnz- ca - ca - cooH._¿> (1, terminal _ 2 thioalkanoyl) X oco (cH2) '-CH = CH »- = L or (b) in the case of the compounds of the formula In the thioalkanoyl group R1COS is in the non-terminal position, a compound of the general formula IV + - (cH3) 3N - CH2 - ca - ca 2 'COOH x oco (cH2) mcH = cn - (cH2) m. - R2 where m and m! each is an integer between 0 and 3 and R2 is a lower alkyl group having 1 to 4 carbon atoms, reacted with a thioacid of the general formula II R1cosH where R1 has the meaning given above, according to the following reaction formula 10 15 20 25 453 191 IS '+. R COSH + (CH) N - CH - CH - CH - COOH -) (I, non-terminal 1 3 3 2 2. Thioalkanoyl) X OCO (CH 2) m - CH = CH - (CH) m. - R2 6. Pharmaceutical composition with mucolytic and antitussive activities, characterized in that it comprises a therapeutically effective amount of at least one of the thio-carnitines of the general formula I + (CH3) 3N - CH2 - 1H - CH2 - COOH X OCO -'R-SCO-R1 where R is a straight alkenyl group having 2 to 6 carbon atoms or a branched alkylene group having 3 to 6 carbon atoms R1 a straight or branched lower alkyl group having 1 to 4 carbon atoms and X- a halide ion in claim 1, as active ingredient and a pharmaceutically acceptable excipient.